The safety and efficacy of a therapeutic agent is a function of (i) its intrinsic biological activity and (ii) the biodistribution achieved after its administration. Many potentially useful therapeutic agents possess a biochemical activity ameliorating a particular pathological condition, but the presence of the agent in normal, nonpathological tissue results in deleterious effects that prevent the use of the agent. Damage to a normally functioning kidney, bone marrow, liver tissue or other organ may limit the use of therapeutic agents with established antiviral activity, or agents with established anti-cancer activity. There is a need for new compounds to target therapeutic agents to the specific cells that are the source of some pathological condition, and to reduce the concentration attained in unaffected, normal tissues. Targeting is the modification of a therapeutic agent so that after injection or oral administration the uptake by a specific population of cells is increased relative to uptake of the unmodified agent. By targeting compounds with established and beneficial biological activity to specific tissues, compounds whose use is currently limited by side effects might become safe and efficacious drugs. A therapeutic agent is a compound administered with the intent of changing in a beneficial manner some physiological function. Therapeutic agents include radioprotective agents, chemoprotective agents, antiviral agents, antibodies, enzymes, and peptides.
One method of targeting therapeutic agents to specific cells involves attaching them to carrier molecules recognized by receptors performing receptor mediated endocytosis. Of particular interest is targeting via the asialoglycoprotein receptor of hepatocytes. This receptor is present in high levels on normal hepatocytes but in lower levels or not at all on transformed hepatocytes (hepatoma cells). Diagnostic and therapeutic agents have been attached to asialoglycoprotein carriers and neoglycoprotein carriers recognized by the asialoglycoprotein receptor and targeted to the cells, see Table II of Meijer and van der Sullies, Pharm. Res. (1989) 6:105-118 and Ranade, J. Clin. Pharmacol. (1989) 29:685-694. Molecules recognizing the asialoglycoprotein receptor are most often either asialoglycoproteins or neoglycoproteins. Asialoglycoproteins are formed by removing the sialic acid of glycoproteins and exposing galactose residues. Neoglycoproteins are formed by attaching multiple galactose residues to non-glycoproteins such as human albumin.
When attaching diagnostic and therapeutic agents to a receptor-recognizing carrier molecule, targeting can be achieved only if the affinity of the carrier for the receptor is maintained. The differential reactivity of the protein amine and carbohydrate hydroxyl groups of glycoprotein carriers, e.g. asialofetuin, is commonly used to achieve this goal. The highly reactive amine groups of protein lysine residues are selectively modified, while the hydroxyl groups of carbohydrate are left intact and continue to recognize the receptor. Examples of this strategy are given in Van der Sluijs et al. (above) and in "Liver Diseases, Targeted Diagnosis and Therapy Using Specific Receptors and Ligands" (1991) Ed. G. Y. Wu and C. H. Wu, Marcel Dekker Inc. pp. 235-264. In contrast, a polysaccharide such as arabinogalactan offers no polypeptide amino groups distal from the receptor binding site that can be modified for the purposes of retaining the asialoglucoprotein receptor binding activity. In spite of the obvious strategy for modification of glycoproteins with retention of receptor binding activity, their use for targeted, parenteral pharmaceuticals is subject to several problems.
(i) Glycoproteins are prepared from animal cells and insuring noncontamination with human infective viral pathogens is a major issue.
(ii) Glycoproteins will not generally tolerate organic solvents during conjugate synthesis, because such solvents frequently lead to a loss of biological activity and denaturation.
(iii) Glycoproteins can be toxic and/or antigenic.
(iv) Glycoproteins in their native form, e.g. fetuin, do not afford galactose resides and must be desialylated to produce a carrier which interacts with the receptor.
Arabinogalactans are a class of polysaccharides obtained from the cell walls of many species of trees and other plants. A common, commercially available source of arabinogalactan is the American Western larch (Larix occidentalis). Arabinogalactan from this source is used as a binder, emulsifier or stabilizer in foods. It consists of a largely 1-3 linked D-galactose backbone with 1-6 linked branch chains of L-arabinoses and D-galactoses at practically every residue on the backbone. In larch arabinogalactans the ratio of galactose to arabinose is between 5 to 1 and 10 to 1, while arabinogalactans from plant sources in general range from about 1 to 4 to about 10 to 1 [Clarke, A. E., Anderson, R. L., Stone, B. A.; Phytochemistry (1979) 18: 521-40]. Like many polysaccharides, arabinogalactans have different molecular weights with values of about 1-2 million to about 10,000 daltons [Blake, J. D., Clarke, M. L., Jansson, P. E.; Carbohydr Res (1983) 115: 265-272] having been reported. It has been shown that L-arabinose and D-galactose interact with the asialoglycoprotein receptor while common monosaccharides like glucose or mannose do not [Lee, Haekyung, Kelm, Sorge, Teruo, Yoshino, Schauer, Roland Biol. Chem., Hoppe-Seyler (1988) 369, 705-714].
Some derivatives of arabinogalactan have been previously prepared. Graft copolymers have been used in paper manufacturing [SU1285094] and in soil treatments [JP1051198]. Arabinogalactan sulfate has been used to form salts with drugs to influence drug absorption and prolong drug action [US4609640]. Acidic forms of arabinogalactan occur naturally having a composition which includes uranic acid [Clarke, A. E., Anderson, R. L., Stone, B. A., Phytochemistry (1979) 18, 521-40], and have also been prepared from arabinogalactan [JP60219202]. Derivatives of arabinogalactan with substituent alkyl, allyl cyano, halo or amino groups, and conjugates with organic acids and enzyme protein have been disclosed, wherein the carbohydrate is used as a carrier, adsorbent or resin [JP60219201]. In some cases arabinogalactan has been highly derivatized in a manner likely to destroy its interaction with the asialoglycoprotein receptor. For example, in some cases as many as 50% of the hydroxyl groups of arabinogalactan have been modified [JP60219201], but the affinity, or lack thereof, of arabinogalactan derivatives for the asialoglycoprotein receptor has not been studied.